Myocardial infarction (MI) results in cardiomyocyte death in the infarct zone followed by scar formation and pathological remodelling of the ventricle. Myocardium does not regenerate and viable tissue remaining is often insufficient to maintain adequate cardiac output. Heart transplants are often not an available or appropriate option. Thus, there is a pressing need for alternative interventions, such as cell replacement therapy.
The first evidence that cell replacement therapy may be a viable intervention for the treatment of MI came from animal studies showing that injection of fetal or neonatal cardiomyocytes (CM) improved left ventricular function and ventricle thickness in the post-MI setting. Injected CM integrated with host CM through gap junctions and intercalated discs. Yet one of the main limitations of these studies and a most probable reason for the incomplete functional recovery was the massive death of injected myocytes.
In most myocardial cell injection studies, the cells were suspended in a liquid such as saline or culture medium followed by intramyocardial or coronary injection. The main challenges associated with this procedure were poor survival of the injected cells and washout from the injection site. According to some estimates, 90% of cells delivered through a needle leaked out of the injection site. In addition, a significant number of cells (˜90%) died within days after injection.
Thus, developing improved delivery and localization methods (e.g. hydrogels) and effective anti-cell death strategies could significantly improve effectiveness of cell injection procedures. Towards that goal, Kofidis et al. (Balsam, Wagers et al. 2004; Kofidis, Lebl et al. 2005), reported that injection of Matrigel or Matrigel plus embryonic stem cells (ESC) into infarcted rat hearts resulted in structural stabilization, prevented wall thinning and improved fractional shortening. Chirstman et al. (Christman, Fok et al. 2004; Christman, Vardanian et al. 2004) demonstrated that injection of skeletal myoblasts into myocardial infarcts using fibrin matrix increased cell localization within the infarct after five weeks, reduced infarct size and increased vascularization without causing a marked inflammatory response or foreign body reaction. Similarly, Ryu Hee et al (Ryu, Kim et al. 2005) found that injection of bone marrow mononuclear cells into cryoinjured rat myocardium using fibrin matrix increased the amount of viable tissue, improved microvessel formation and reduced the amount of fibrous tissue in comparison to the injection of cells in culture medium or culture medium alone. (Laflamme, Gold et al. 2005) Laflamme and Murry demonstrated that using Matrigel modified with a number of biomolecules to target multiple pathways related to cell survival, significantly increased the grafting of the human ESC-derived CM injected into infracted rat hearts (Laflamme, Chen et al. 2007).
It was also demonstrated that a synthetic material, self-assembling peptide hydrogel, could be utilized for cell injection into the myocardium (Davis, Motion et al. 2005). Upon injection, the peptide formed a nano-fibrous structure that promoted recruitment of endogenous cells expressing endothelial markers, and supported survival of injected CM. The peptide consisted of alternating hydrophilic and hydrophobic domains (AcN-RARADADARARADADA-CNH) (SEQ ID NO: 1) and did not activate integrin signalling. Insulin-like growth factor-1 bound to the self-assembling peptide improved grafting and survival of CM injected into infarcted myocardium (Davis, Hsieh et al. 2006). Photocrosslinkable PEGylated fibrinogen was recently demonstrated to be an excellent substrate for encapsulation and cultivation of CM derived from neonatal rat hearts and human ESC-derived CM. The cells cultivated in these hydrogels were connected to each other via gap junctions and demonstrated significant cross-striations (Shapira-Schweitzer, Habib et al. 2009).
Recent studies collectively indicate that an injection of hydrogel alone, without the reparative cells, may also attenuate pathological remodeling upon myocardial infarction (Landa, Miller et al. 2008; Dobner, Bezuidenhout et al. 2009; Fujimoto, Ma et al. 2009; Leor, Tuvia et al. 2009). It is thought that hydrogels act by changing the ventricular geometry and mechanics, thus reducing elevated local wall stresses that have been implicated in pathological remodeling (Wall, Walker et al. 2006). Finite element modeling of wall stresses indicated that upon injection of the material of elastic modulus 10−20 kPa in the infarct, injection improved ejection fraction and the stroke volume/end-diastolic volume relationship. In addition, injections of the material in the border zone decreased end-systolic fiber stress proportionally to the volume and the stiffness of the injected material.
Angiopoietin 1 (ang1) is known to preserve cardiac function post-MI (Siddiqui, Blomberg et al. 2003) (Zhou, Ma et al. 2005) in animal models. Ang1 interacts directly with CM via integrins to promote adhesion, survival, cardioprotective signalling (akt/MAPK) and prevent induced apoptosis (Dallabrida, Ismail et al. 2005). CMs survived better on immobilized ang1 then on most other matrices present in the heart. Ang1 lacks known integrin-binding motifs, but the novel site QHREDGS (SEQ ID NO: 2) was identified (Dallabrida, Ismail et al. 2005). Further, ang1/integrin interactions attenuated cardiac hypertrophy in vivo (Dallabrida, Ismail et al. 2008).
Low survival rate during culture and passaging of human induced pluripotent stem (iPS) cells presents a major obstacle in research; especially hindering further manipulations during induction and differentiation processes to obtain functional cells for regenerative therapies. Recent reports suggest that the addition of Y-27632, a selective inhibitor of p160-Rho-associated coiled-coil kinase (ROCK), to culture media permits survival of dissociated human ES cells during passaging without compromising pluripotency or differentiation potential. However, because Rho/ROCK can activate different signaling cascades depending on cell type and environmental context, as well as contribute to changes in the cytoskeleton, it is possible that treatment of human iPS cells with its inhibitor, Y-27632, may result in less than favorable conditions for the subsequent differentiation of these cells.
It would be desirable, thus, to develop survival promoting means effective to treat conditions involving cell death and apoptosis.